Microbiome, Transcriptome, and Metabolomic Analyses Revealed the Mechanism of Immune Response to Diarrhea in Rabbits Fed Antibiotic-Free Diets

Abstract

In this study, diarrhea was induced in rabbits by feeding them antibiotic-free feed. The gut provides important defense against the barriers of the body, of which the duodenum is an important part to help digest food and absorb nutrients. However, the mechanisms underlying the roles of the gut microbiome and fecal metabolome in rabbit diarrhea caused by feeding an antibiotic-free diet have not been characterized. Recently, only a single study has been conducted to further characterize the antibiotic-free feed additives that caused diarrhea in weaned rabbits. The multi-omics techniques, including 16S rRNA sequencing, transcriptome sequencing, and LC-MS analysis, were combined to analyze the gut microbial compositions and functions. They also determined the fecal metabolomic profiles of diarrhea in rabbits caused by feeding antibiotic-free feed. The results showed that the liver, duodenal, and sacculus rotundus tissues of diarrhea rabbits were diseased, the composition of intestinal microbes was significantly changed, the diversity of intestinal microbes was decreased, and the distribution of intestinal microbe groups was changed. Functional analysis based on the cluster of GO and KEGG annotations suggested that two functional GO categories belonged to the metabolism cluster, and five KEGG pathways related to the metabolic pathways were significantly enriched in diarrhea rabbits. Moreover, real-time quantitative PCR (RT-qPCR) was used to verify the significant expression of genes related to diarrhea. Metabolomics profiling identified 432 significantly differently abundant metabolites in diarrhea rabbits, including amino acids and their derivatives. These amino acids were enriched in the tryptophan metabolic pathway. In addition, the functional correlation analysis showed that some altered gut microbiota families, such as Parasutterella, significantly correlated with alterations in fecal metabolites. Collectively, the results suggested that altered gut microbiota was associated with diarrhea caused by antibiotic-free feed additives in weaned rabbit pathogenesis.

Genome‐wide association studies for diarrhoea outcomes identified genomic regions affecting resistance to a severe enteropathy in suckling rabbits

“…In the field conditions in which we nested our study, it was not possible to obtain specimens from all involved suckling rabbits (as the healthy animals could not be sampled); therefore, we opted for a first characterization of the diarrhoea microbiota of the deceased rabbits, sampled from different litters, that may correspond to the final stage of the disease. Despite the observed heterogeneity of the microbiota profiles across samples, also reported within experimental design in other studies (Chen et al., 2022; Wang, Fan, et al., 2022), the most consistent bacterial family and genus across all investigated specimens was Clostridiaceae (24%–57% of the reads of the 16S rRNA gene profiles) and Clostridium (24%–66% of the reads of the same profile), respectively, which might include pathogenic agents or might provide a peculiar signature of the resulted disease status. The genus Clostridium has been reported by the studies of Velasco‐Galilea et al.…”

“…Therefore, mainly due to the heterogeneity and complexity of these types of disorders in rabbits and, in turn, the complications of the experimental procedures that should be designed, none of the studies that thus far investigated, at the rabbit genome level, DNA markers associated with this pathogenic condition, also attempted to obtain a precise characterization of the putative biological components (e.g., microbiota, viroma) involved as causative factors or derived consequence of the disorder. Only few studies reported information on the microbiota characterization of diarrhoea in rabbits linked to different feeding strategies in specific experimental designs (Chen et al., 2022; Wang, Fan, et al., 2022).…”

Integration of Non-Coding RNA and mRNA Profiles Reveals the Mechanisms of Rumen Development Induced by Different Types of Diet in Calves

“…We performed qRT-PCR in a CFX96 Real-Time PCR Detection system (Bio-RadCo., Hercules, CA, USA) and detected RNA expression using SYBR Green Real-Time PCR Master Mix (Takara Co., Dalian, China). The relative expression levels were calculated using the 2 −∆∆Ct method and normalized to those of the reference genes ACTB and U6 [45,46].…”

Seasonal shift in gut microbiome diversity in wild Sichuan takin (Budorcas tibetanus) and environmental adaptation

“…Therefore, we speculate that specific pathways must have been activated/deactivated during seasonal change, leading to up- or down-regulation in gene transcription and hence protein levels, ultimately leading to physiological changes in the animals that help them adapt to the environment. To follow up on this association between metabolic pathways and seasonal change, future transcriptomic and metabolomic studies focusing on specific mRNA, proteins and other small molecules as a more robust evaluation of metabolic function for verifying the predicted pathways are warranted [32] , [33] , [34] .…”